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1.
Int J Mol Sci ; 24(22)2023 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-38003578

RESUMO

Drought stress profoundly affects plant growth and development, posing a significant challenge that is extensively researched in the field. Thioredoxins (TRXs), small proteins central to redox processes, are crucial to managing both abiotic and biotic stresses. In this research, the VyTRXy gene, cloned from wild Yanshan grapes, was validated as a functional TRX through enzyme activity assays. VyTRXy was found to bolster photosynthesis, augment levels of osmotic regulators, stimulate antioxidant enzyme activities, and strengthen drought resilience in transgenic plants. These enhancements were evidenced by higher survival rates, optimized photosynthetic metrics, increased proline levels, augmented chlorophyll concentration, reduced electrolyte leakage, and decreased malondialdehyde and hydrogen peroxide (H2O2) levels. Furthermore, there was a surge in the activities of enzymes such as catalase, ascorbate peroxidase, glutathione peroxidase, dehydroascorbate reductase, and glutathione reductase, along with an increased expression of TRX peroxidase. Notably, under drought stress, there was a marked elevation in the expression of stress-responsive genes, including the adversity stress-inducible expression gene (NtRD29A) and DRE-binding protein (NtDREB), in transgenic tobacco. This investigation is pivotal in the quest for drought-resistant grapevine varieties and provides significant insights into the molecular functionality of VyTRXy in enhancing grapevine drought tolerance.


Assuntos
Antioxidantes , Resistência à Seca , Antioxidantes/metabolismo , Peróxido de Hidrogênio/metabolismo , Fotossíntese , Estresse Fisiológico/genética , Plantas Geneticamente Modificadas/metabolismo , Secas , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Espécies Reativas de Oxigênio/metabolismo
2.
Front Plant Sci ; 14: 1173857, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37223813

RESUMO

Salt stress is an important factor which may negatively affect plant growth and development. High concentrations of Na+ ions can destroy the ion balance in plant somatic cells, as well as destroying cell membranes and forming a large number of reactive oxygen species (ROS) and other damage mechanisms. However, plants have evolved numerous defense mechanisms in response to the damages caused by salt stress conditions. Grape (Vitis vinifera L.), a type of economic crop, is widely planted throughout the world. It has been found that salt stress is an important factor affecting the quality and growth of grape crops. In this study, a high-throughput sequencing method was used to identify the differentially expressed miRNAs and mRNAs in grapes as responses to salt stress. A total of 7,856 differentially expressed genes under the salt stress conditions were successfully identified, of which 3,504 genes were observed to have up-regulated expressions and 4,352 genes had down-regulated expressions. In addition, this study also identified 3,027 miRNAs from the sequencing data using bowtie and mireap software. Among those, 174 were found to be highly conserved, and the remaining miRNAs were less conserved. In order to analyze the expression levels of those miRNAs under salt stress conditions, a TPM algorithm and DESeq software were utilized to screen the differentially expressed miRNAs among different treatments. Subsequently, a total of thirty-nine differentially expressed miRNAs were identified, of which fourteen were observed to be up-regulated miRNAs and twenty-five were down-regulated under the salt stress conditions. A regulatory network was built in order to examine the responses of grape plants to salt stress, with the goal of laying a solid foundation for revealing the molecular mechanism of grape in responses to salt stress.

3.
Plant Physiol Biochem ; 200: 107754, 2023 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-37236064

RESUMO

Cadmium (Cd) contamination in agricultural soils has become a serious worldwide environmental problem threatening crop production and human health. Hydrogen peroxide (H2O2) is a critical second messenger in plant response to Cd exposure. However, its role in Cd accumulation in various organs of plants and the mechanistic basis of this regulation remains to be elucidated. In this study, we used electrophysiological and molecular approaches to understand how H2O2 regulates Cd uptake and translocation in rice plants. Our results showed that the pretreatment of H2O2 significantly reduced Cd uptake by rice roots, which was associated with the downregulation of OsNRAMP1 and OsNRAMP5. On the other hand, H2O2 promoted the root-to-shoot translocation of Cd, which might be attributed to the upregulation of OsHMA2 critical for Cd2+ phloem loading and the downregulation of OsHMA3 involved in the vacuolar compartmentalization of Cd2+, leading to the increased Cd accumulation in rice shoots. Furthermore, such regulatory effects of H2O2 on Cd uptake and translocation were notably amplified by the elevated level of exogenous calcium (Ca). Collectively, our results suggest that H2O2 can inhibit Cd uptake but increase root to shoot translocation through modulating the transcriptional levels of genes encoding Cd transporters, furthermore, application of Ca can amplify this effect. These findings will broaden our understanding of the regulatory mechanisms of Cd transport in rice plants and provide theoretical foundation for breeding rice for low Cd accumulation.


Assuntos
Oryza , Poluentes do Solo , Humanos , Cádmio/metabolismo , Peróxido de Hidrogênio/farmacologia , Oryza/metabolismo , Melhoramento Vegetal , Transporte Biológico , Proteínas de Membrana Transportadoras , Raízes de Plantas/metabolismo , Poluentes do Solo/análise
4.
Int J Mol Sci ; 23(23)2022 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-36499201

RESUMO

Phenolic extracts from berry seeds have been extensively studied for their health benefits. However, few studies have been conducted on the effects of phenolic extracts from Vitis L. canes and berry stems. The Chinese spine grape (V. davidii Foex) is an important and widely distributed wild species of Vitis L. The present study explored the metabolomic profile and evaluated the antioxidant activity of phenolic compounds in extracts from V. davidii Foex. canes and stems, with a focus on their role in preventing DNA damage caused by free radicals and inhibiting the growth of breast (MCF-7) and cervical (HeLa) cancer cells. Total phenolic compounds in the dried berry stems of spine grapes were higher than that in vine canes. Analysis of the extracts showed that proanthocyanins, epicatechin, catechin, and phenolic acid were the main phenolic compounds in V. davidii Foex, but in higher quantities in berry stems than in vine canes. However, trans-resveratrol and kaempferol 3-O-glucoside were present in the vine canes but not in the berry stems. Antioxidant analysis by FRAP and ABTS showed that extracts from berry stems and vine canes had a higher antioxidant activity than thinned young fruit shoots before flowering, leaves, peel, pulp, and seeds in V. davidii Foex. Moreover, the antioxidant activity of extracts from berry stems was higher than that in other grape species, except for muscadine. In vitro analyses further showed that the extracts significantly increased H2O2 scavenging ability and conferred a protective effect against DNA damage. Furthermore, a low concentration of phenolic compounds in extracts from the vine canes and berry stems of spine grapes inhibited the proliferation of the MCF-7 and Hela cancer cells. These research results provided some important useful information for the exploitation of V. davidii Foex canes and berry stems and indicated that canes and stems of V. davidii Foex had good antioxidant properties, anticancer activity and prevented DNA damage, providing evidence for medical utilization of V. davidii Foex.


Assuntos
Catequina , Vitis , Vitis/genética , Antioxidantes/farmacologia , Antioxidantes/análise , Peróxido de Hidrogênio , Fenóis/farmacologia , Fenóis/análise , Frutas/química , Extratos Vegetais/farmacologia , Extratos Vegetais/análise
5.
Front Plant Sci ; 13: 1009747, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36311118

RESUMO

Temperature is one of the most important factors regarding fruit postharvest, however its effects in the strawberry fruits quality in postharvest remains to be evaluated. In this study, the effects of cold and heat storage temperature on fruit quality of 'Benihoppe' strawberry were performed. The results showed that different temperatures could affect the metabolism of hormone, anthocyanin, reactive oxygen species (ROS), and transcription level of responsive factors. The synthesis of terpenoids, amino acids, and phenylpropanoids in strawberries were also changed under different temperatures, which finally changed the quality characteristics of the fruit. We found HSF20 (YZ1)-overexpressed fruits were sensitive to cold and heat conditions but CBF/NF-Y (YZ9)-overexpressed fruits promoted coloring under cold treatment. This study clarified the effect of postharvest cooling and heat treatments on quality and transcriptional mechanism of strawberries fruits. Moreover, these results provided an experimental basis for further research on improving the quality of strawberry berries during postharvest periods.

6.
Plant Signal Behav ; 17(1): 2000791, 2022 12 31.
Artigo em Inglês | MEDLINE | ID: mdl-35152834

RESUMO

Necrosis and ethylene-inducing peptide 1 (Nep1) -like proteins (NLP) are secreted by multiple taxonomically unrelated plant pathogens (bacteria, fungi, and oomycete) and are best known for inducing cell death and immune responses in dicotyledonous plants. A group of putative NLP genes from obligate biotrophic oomycete Plasmopara viticola were predicted by RNA-Seq in our previous study, but their activity has not been established. Therefore, we analyzed the P. viticola NLP (PvNLP) family and identified seven PvNLP genes. They all belong to type 1 NLP genes and form a P. viticola-specific cluster when compared with other pathogen NLP genes. The expression of PvNLPs was induced during early infection process and the expression patterns could be categorized into two groups. Agrobacterium tumefaciens-mediated transient expression assays revealed that only PvNLP7 was cytotoxic and could induce Phytophthora capsici resistance in Nicotiana benthamiana. Functional analysis showed that PvNLP4, PvNLP5, PvNLP7, and PvNLP10 significantly improved disease resistance of Arabidopsis thaliana to Hyaloperonospora arabidopsidis. Moreover, the four genes caused an inhibition of plant growth which is typically associated with enhanced immunity when over-expressed in Arabidopsis. Further research found that PvNLP7 could activate the expression of defense-related genes and its conserved NPP1 domain was critical for cell death- and immunity-inducing activity. This record of NLP genes from P. viticola showed a functional diversification, laying a foundation for further study on pathogenic mechanism of the devastating pathogen.


Assuntos
Arabidopsis , Oomicetos , Phytophthora , Arabidopsis/metabolismo , Resistência à Doença , Oomicetos/fisiologia , Phytophthora/fisiologia , Doenças das Plantas/microbiologia , Proteínas/química , /metabolismo
7.
Trends Plant Sci ; 27(9): 890-907, 2022 09.
Artigo em Inglês | MEDLINE | ID: mdl-35165036

RESUMO

Gene editing using clustered regularly interspaced short palindromic repeat/CRISPR-associated proteins (CRISPR/Cas) has revolutionized biotechnology and provides genetic tools for medicine and life sciences. However, the application of this technology to miRNAs, with the function as negative gene regulators, has not been extensively reviewed in plants. Here, we summarize the evolution, biogenesis, and structure of miRNAs, as well as their interactions with mRNAs and computational models for predicting target genes. In addition, we review current advances in CRISPR/Cas for functional analysis and for modulating miRNA genes in plants. Extending our knowledge of miRNAs and their manipulation with CRISPR will provide fundamental understanding of the functions of plant miRNAs and facilitate more sustainable and publicly acceptable genetic engineering of crops.


Assuntos
Sistemas CRISPR-Cas , MicroRNAs , Sistemas CRISPR-Cas/genética , Produtos Agrícolas/genética , Evolução Molecular , Edição de Genes , MicroRNAs/genética
8.
Front Plant Sci ; 13: 1061520, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36643293

RESUMO

Entomopathogen-based biocontrol is crucial for blocking the transmission of vector-borne diseases; however, few cross-latitudinal investigations of entomopathogens have been reported for vectors transmitting woody plant diseases in forest ecosystems. The pine sawyer beetle Monochamus alternatus is an important wood borer and a major vector transmitting pine wilt disease, facilitating invasion of the pinewood nematode Bursaphelenchus xylophilus (PWN) in China. Due to the limited geographical breadth of sampling regions, species diversity of fungal associates (especially entomopathogenic fungi) on M. alternatus adults and their potential ecological functions have been markedly underestimated. In this study, through traditional fungal isolation with morphological and molecular identification, 640 fungal strains (affiliated with 15 genera and 39 species) were isolated from 81 beetle cadavers covered by mycelia or those symptomatically alive across five regional populations of this pest in southern China. Multivariate analyses revealed significant differences in the fungal community composition among geographical populations of M. alternatus, presenting regionalized characteristics, whereas no significant differences were found in fungal composition between beetle genders or among body positions. Four region-representative fungi, namely, Lecanicillium attenuatum (Zhejiang), Aspergillus austwickii (Sichuan), Scopulariopsis alboflavescens (Fujian), and A. ruber (Guangxi), as well as the three fungal species Beauveria bassiana, Penicillium citrinum, and Trichoderma dorotheae, showed significantly stronger entomopathogenic activities than other fungi. Additionally, insect-parasitic entomopathogenic fungi (A. austwickii, B. bassiana, L. attenuatum, and S. alboflavescens) exhibited less to no obvious phytopathogenic activities on the host pine Pinus massoniana, whereas P. citrinum, Purpureocillium lilacinum, and certain species of Fusarium spp.-isolated from M. alternatus body surfaces-exhibited remarkably higher phytopathogenicity. Our results provide a broader view of the entomopathogenic fungal community on the vector beetle M. alternatus, some of which are reported for the first time on Monochamus spp. in China. Moreover, this beetle might be more highly-risk in pine forests than previously considered, as a potential multi-pathogen vector of both PWN and phytopathogenic fungi.

9.
Front Plant Sci ; 12: 761713, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-35058944

RESUMO

Flowering is the key process for the sexual reproduction in seed plants. In gramineous crops, the process of flowering, which includes the actions of both glume opening and glume closing, is directly driven by the swelling and withering of lodicules due to the water flow into and out of lodicule cells. All these processes are considered to be controlled by aquaporins, which are the essential transmembrane proteins that facilitate the transport of water and other small molecules across the biological membranes. In the present study, the evolution of aquaporins and their contribution to flowering process in plants were investigated via an integration of genome-wide analysis and gene expression profiling. Across the barley genome, we found that HvTIP1;1, HvTIP1;2, HvTIP2;3, and HvPIP2;1 were the predominant aquaporin genes in lodicules and significantly upregulated in responding to glume opening and closing, suggesting the importance of them in the flowering process of barley. Likewise, the putative homologs of the above four aquaporin genes were also abundantly expressed in lodicules of the other monocots like rice and maize and in petals of eudicots like cotton, tobacco, and tomato. Furthermore, all of them were mostly upregulated in responding to the process of floret opening, indicating a conserved function of these aquaporin proteins in plant flowering. The phylogenetic analysis based on the OneKP database revealed that the homologs of TIP1;1, TIP1;2, TIP2;3, and PIP2;1 were highly conserved during the evolution, especially in the angiosperm species, in line with their conserved function in controlling the flowering process. Taken together, it could be concluded that the highly evolutionary conservation of TIP1;1, TIP1;2, TIP2;3 and PIP2;1 plays important roles in the flowering process for both monocots and eudicots.

10.
Electrophoresis ; 41(21-22): 1832-1842, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32436592

RESUMO

Dynamic pH barrage junction focusing in CE enables effective signal enhancement, quantitative capture efficiencies, and straightforward optimization. The method is a technical variant of dynamic pH junction focusing. CE separation with dynamic pH barrage junction focusing is compatible with both optical and mass spectrometric detection. We developed a CE-MS/MS method using hydrophilic polyethyleneimine-coated capillaries and validated it for the qualitative analysis of amino acids, peptides, and tryptic peptides of digested monoclonal antibodies. The S/N of extracted ion electropherograms of zwitterionic analytes were enhanced by approximately two orders of magnitude with a tradeoff of a shortened separation window. Online focusing improved the MS signal intensity of a diluted antibody digest, enabling more precursor ions to be analyzed with subsequent tandem mass spectrometric identification. It also broadened the concentration range of protein digest samples for which adequate sequence coverage data can be obtained. With only 0.9 ng of digested infliximab sample loaded into the capillary, 76% and 100% sequence coverage was realized for antibody heavy and light chains, respectively, after online focusing. Full coverage was achieved with 9 ng of injected digest.


Assuntos
Aminoácidos/análise , Anticorpos Monoclonais/análise , Eletroforese Capilar/métodos , Peptídeos/análise , Espectrometria de Massas em Tandem/métodos , Aminoácidos/química , Anticorpos Monoclonais/química , Eletroforese Capilar/instrumentação , Desenho de Equipamento , Concentração de Íons de Hidrogênio , Peptídeos/química , Espectrometria de Massas em Tandem/instrumentação , Tripsina
11.
J Proteome Res ; 19(7): 2700-2707, 2020 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-32338916

RESUMO

Mass spectrometry is a powerful tool for de novo sequencing of novel proteins. Recent efforts in this area have mainly focused on liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Here, we present an alternative method, capillary electrophoresis tandem mass spectrometry (CE-MS/MS), for sequencing novel monoclonal antibodies. Using less than 200 ng in total of tryptic digest sample in a triplicated measurement, CE-MS/MS with pH-mediated focusing successfully sequenced mAb infliximab with 100% sequence coverage and 100% accuracy for the light chain and 96% coverage and 93% accuracy for the heavy chain. It was also demonstrated that CE-MS/MS gives comparable results, and in some cases, even better results, as compared to LC-MS/MS when used as a standalone technique. A combined workflow using both CE-MS/MS and LC-MS/MS was also used to sequence a novel antibody, anti-CD-176, resulting in the first proposed sequence for this mAb.


Assuntos
Anticorpos Monoclonais , Espectrometria de Massas em Tandem , Cromatografia Líquida , Eletroforese Capilar , Análise de Sequência
12.
Electrophoresis ; 41(5-6): 370-378, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31994203

RESUMO

A high organic content CE-MS/MS (HOCE-MS/MS) method was developed for the proteomic analysis of envelope proteins extracted from spinach leaves. Separation was performed in a 1-m long hydroxypropyl cellulose coated capillary, using 8% (v/v) formic acid in 70% (v/v) methanol and 22% water as the BGE. A flow-through microvial interface was used to couple the CE system with an Orbitrap Fusion Lumos mass spectrometer, and field-amplified sample stacking was used to improve the concentration sensitivity. Using this optimized method, 3579 peptides and 1141 proteins were identified using the Proteome Discoverer software with a 1% false discovery rate at the protein level. Relative to conventional aqueous CE, HOCE-MS did a better job of discovering hydrophobic peptides and provided more peptide and protein identifications. Relative to nano-LC-MS, it achieved comparable peptide and protein identification performance and detected peptides not identified by LC-MS: of the full set of peptides identified using the two techniques, 19% were identified only using HOCE-MS. It also outperformed nano-LC-MS with respect to the detection of low molecular weight peptides.


Assuntos
Proteínas de Cloroplastos , Eletroforese Capilar/métodos , Proteômica/métodos , Spinacia oleracea/química , Espectrometria de Massas em Tandem/métodos , Proteínas de Cloroplastos/análise , Proteínas de Cloroplastos/química , Proteínas de Cloroplastos/isolamento & purificação , Cloroplastos/química , Interações Hidrofóbicas e Hidrofílicas , Folhas de Planta/química , Proteoma/análise , Proteoma/química
13.
Mitochondrial DNA B Resour ; 5(3): 2855-2856, 2020 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-33457976

RESUMO

Vitis champinii is a grapevine rootstock species and widely used in vineyards and in rootstock breeding programs for regions with high nematode populations or saline soils. Here, the complete chloroplast genome of V. champinii was reported. The length of the chloroplast genome was 160,657 bp with a large single copy region of 89,217 bp, a small single copy region of 19,504 bp and two separated inverted regions of 51,936 bp, respectively. In total, 130 unique genes were identified of this genome, including 85 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. Phylogenetic analysis indicates that V. champinii is closely related to Vitis acerifolia.

14.
Mitochondrial DNA B Resour ; 5(3): 3050-3051, 2020 Jul 29.
Artigo em Inglês | MEDLINE | ID: mdl-33458054

RESUMO

Vitis berlandieri, a species of grape native to the southern North America, is known for good tolerance against soils with a high content of lime and was almost used for rootstock breeding. Here, we report the complete chloroplast genome of V. berlandieri. The chloroplast genome was 161,028 bp in length, harboring a large single-copy region (89,228 bp) and a small single-copy region (19,028 bp) separated by two inverted repeat regions. A total of 130 unique genes were identified from this genome, including 85 protein-coding genes (PCGs), 37 tRNA genes, and 8 rRNA genes. Chloroplast phylogenetic analysis revealed that V. berlandieri is closely related to V. cordifolia.

15.
Electrophoresis ; 39(9-10): 1216-1221, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-28990192

RESUMO

A nonaqueous capillary electrophoresis mass spectrometry (NACE-MS) method was developed to separate and determine highly hydrophobic temporin peptides. The nonaqueous background electrolyte solution was a mixture of 20% acetonitrile, 78% methanol and 2% formic acid, with 20 mM ammonium formate. The separation of six peptides was completed within 12 min. The CE system was connected to a triple quadrupole mass spectrometer operating in MRM mode using a chemical modifier solution of 2 mM ammonium formate in ethanol with the flow through microvial interface. The mass spectrometer offered a second dimension of separation for peptides having identical migration times but different structures. The new method represents the first system capable of reliably determining hydrophobic peptides without using reversed phase liquid chromatography mass spectrometry.


Assuntos
Eletroforese Capilar/métodos , Peptídeos/análise , Peptídeos/isolamento & purificação , Espectrometria de Massas por Ionização por Electrospray/métodos , Concentração de Íons de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas
16.
BMC Ophthalmol ; 17(1): 191, 2017 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-29029601

RESUMO

BACKGROUND: To determine the prevalence and associated factors for myopia and high myopia among older population in a rural community in Eastern China. METHODS: A community-based, cross-sectional survey was conducted in the Weitang town located in Suzhou, an urban metropolis in East China. A total of 5613 Chinese residents aged 60 years and older were invited to complete a questionnaire and participated in a detailed eye examination,including measurements of visual acuity and refractive error using autorefraction and subjective refraction. Myopia and high myopia was defined as SE < -0.5 diopters (D) and < -5.0 D, respectively. RESULTS: Among the 5613 participating individuals, 4795 (85.4%) complete refraction data of phakic right eye was included for analysis. The age-adjusted prevalence was 21.1% (95% confidence interval [CI], 19.9-22.2) for myopia and 2.5% (95% CI, 2.1-2.9) for high myopia. The prevalence of myopia tended to increase significantly with age(p < 0.001),and women had a higher rate of myopia than men (p < 0.001). According to multivariate logistic regression analysis, adults who were older (odds ration[OR]:1.05; 95% CI:1.04-1.07), spent more time for sleeping at night (OR:1.12;95% CI: 1.06-1.18),or had cataract (OR:1.60;95% CI:1.36-1.88) and family history of myopia (OR:1.47;95% CI:1.23-1.77), are more susceptible to myopia (p < 0.001). People who had older age, family history, cataract and specially longer night-time sleep duration, would have a higher risk of myopia. CONCLUSION: Myopia and high myopia among rural old adult population in Eastern China presents common. The current literature unanticipated suggests that there was a positive significant association between prevalence of myopia and night-time sleep duration among adult. Our data provide some evidence of this relationship and highlight the need for larger studies to further investigate this relationship longitudinally and explore mechanism therein.


Assuntos
Miopia/epidemiologia , Distribuição por Idade , Idoso , Idoso de 80 Anos ou mais , Povo Asiático/etnologia , China/epidemiologia , Estudos Transversais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Miopia Degenerativa/epidemiologia , Prevalência , Fatores de Risco , População Rural/estatística & dados numéricos , Distribuição por Sexo , Inquéritos e Questionários , População Urbana/estatística & dados numéricos , Acuidade Visual
17.
Anal Chem ; 89(3): 1411-1415, 2017 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-28208307

RESUMO

Nonaqueous capillary electrophoresis (NACE) is very well suited for online coupling with mass spectrometry due to the relatively high volatility and low surface tension of most organic solvents. Here we present a quantitative NACE-ESI-MS/MS method for separating and determining physcion, chrysophanol, and aloe-emodin in rhubarb. Dantron was used as an internal standard to ensure accuracy and reproducibility in quantitative analyses. Parameters including the pH, background electrolyte (BGE) composition, flow-through microvial chemical modifier solution composition, and modifier solution flow rate were carefully optimized. The developed method was validated by assessing its precision, LODs, and linear range. The contents of physcion, chrysophanol, and aloe-emodin in rhubarb were determined to be 0.22%, 1.0%, and 0.17%, respectively.


Assuntos
Eletroforese Capilar/métodos , Extratos Vegetais/química , Rheum/química , Espectrometria de Massas por Ionização por Electrospray/métodos , Antraquinonas/análise , Eletrólitos/química , Emodina/análogos & derivados , Emodina/análise , Limite de Detecção , Rheum/metabolismo
18.
Int J Ophthalmol ; 9(11): 1578-1583, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27990359

RESUMO

AIM: To investigate the effect of endothelial progenitor cells (EPCs) labeled by carboxy fluorescein diacetate succinimidyl ester (CFSE) on murine oxygen-induced retinopathy (OIR) by intravitreal transplantation. METHODS: After isolated from human umbilical cord blood mononuclear cells, EPCs were cultivated and then labeled with CFSE in vitro. C57BL/6J mice were placed to 75% hyperoxia chamber from P7 to P12 to establish OIR model. At P12, OIR mice were intravitreally injected with 1 µL suspension contained 2×105 EPCs (EPCs group) or isometric phosphate buffered saline (PBS group). The contralateral eye of each mice received no injection (OIR group). Evans blue angiography and frozen section were examined to track the labeled cells in OIR group at P15 and P19. Using retina paraffin sections and adenosinediphos phatase staining at P12 and P19, the effect of EPCs on OIR mice was evaluated quantitatively and qualitatively. RESULTS: The retinas from EPCs group with less non-perfusion area and fewer peripheral tufts were observed at P19, comparing with that from PBS or OIR group. The retinopathy in EPCs group receded earlier with less non-ganglion cells and neovascular nuclei, together with relatively regular distribution. The counts of the neovascular nuclei at P19 were reduced by 44% or 45%, compared with those of OIR group or PBS group respectively. Three days after EPCs injection, a large number of EPCs appeared in the vitreous cavity and adhered to the retinal surface. While at one week, the cells gathered between the internal plexiform layer and the inner limiting membrane, and some EPCs appeared in retinal vessels. CONCLUSION: EPCs transplantation can participate in the reparative procedure of the neovascularization in OIR.

19.
Braz J Microbiol ; 46(2): 601-11, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-26273280

RESUMO

Deinococcus radiodurans (DR) is an extremophile that is well known for its resistance to radiation, oxidants and desiccation. The gene dr1790 of D. radiodurans was predicted to encode a yellow-related protein. The primary objective of the present study was to characterize the biological function of the DR1790 protein, which is a member of the ancient yellow/major royal jelly (MRJ) protein family, in prokaryotes. Fluorescence labeling demonstrated that the yellow-related protein encoded by dr1790 is a membrane protein. The deletion of the dr1790 gene decreased the cell growth rate and sensitivity to hydrogen peroxide and radiation and increased the membrane permeability of D. radiodurans. Transcript profiling by microarray and RT-PCR analyses of the dr1790 deletion mutant suggested that some genes that are involved in protein secretion and transport were strongly suppressed, while other genes that are involved in protein quality control, such as chaperones and proteases, were induced. In addition, the expression of genes with predicted functions that are involved in antioxidant systems, electron transport, and energy metabolism was significantly altered through the disruption of dr1790. Moreover, the results of proteomic analyses using 2-DE and MS also demonstrated that DR1790 contributed to D. radiodurans survival. Taken together, these results indicate that the DR1790 protein from the ancient yellow protein family plays a pleiotropic role in the survival of prokaryotic cells and contributes to the extraordinary resistance of D. radiodurans against oxidative and radiation stresses.


Assuntos
Deinococcus/genética , Genes Bacterianos , Pleiotropia Genética , Mutagênese Insercional , Proteínas de Bactérias/genética , Membrana Celular/fisiologia , Deinococcus/efeitos dos fármacos , Deinococcus/crescimento & desenvolvimento , Deinococcus/efeitos da radiação , Deleção de Genes , Perfilação da Expressão Gênica , Teste de Complementação Genética , Peróxido de Hidrogênio/toxicidade , Proteínas de Membrana/genética , Análise em Microsséries , Viabilidade Microbiana/efeitos dos fármacos , Viabilidade Microbiana/efeitos da radiação , Permeabilidade , Radiação Ionizante , Reação em Cadeia da Polimerase em Tempo Real
20.
Braz. j. microbiol ; 46(2): 601-611, Apr-Jun/2015. tab, graf
Artigo em Inglês | LILACS | ID: lil-749726

RESUMO

Deinococcus radiodurans (DR) is an extremophile that is well known for its resistance to radiation, oxidants and desiccation. The gene dr1790 of D. radiodurans was predicted to encode a yellow-related protein. The primary objective of the present study was to characterize the biological function of the DR1790 protein, which is a member of the ancient yellow/major royal jelly (MRJ) protein family, in prokaryotes. Fluorescence labeling demonstrated that the yellow-related protein encoded by dr1790 is a membrane protein. The deletion of the dr1790 gene decreased the cell growth rate and sensitivity to hydrogen peroxide and radiation and increased the membrane permeability of D. radiodurans. Transcript profiling by microarray and RT-PCR analyses of the dr1790 deletion mutant suggested that some genes that are involved in protein secretion and transport were strongly suppressed, while other genes that are involved in protein quality control, such as chaperones and proteases, were induced. In addition, the expression of genes with predicted functions that are involved in antioxidant systems, electron transport, and energy metabolism was significantly altered through the disruption of dr1790. Moreover, the results of proteomic analyses using 2-DE and MS also demonstrated that DR1790 contributed to D. radiodurans survival. Taken together, these results indicate that the DR1790 protein from the ancient yellow protein family plays a pleiotropic role in the survival of prokaryotic cells and contributes to the extraordinary resistance of D. radiodurans against oxidative and radiation stresses.


Assuntos
Deinococcus/genética , Genes Bacterianos , Pleiotropia Genética , Mutagênese Insercional , Proteínas de Bactérias/genética , Membrana Celular/fisiologia , Deinococcus/efeitos dos fármacos , Deinococcus/crescimento & desenvolvimento , Deinococcus/efeitos da radiação , Deleção de Genes , Perfilação da Expressão Gênica , Teste de Complementação Genética , Peróxido de Hidrogênio/toxicidade , Análise em Microsséries , Proteínas de Membrana/genética , Viabilidade Microbiana/efeitos dos fármacos , Viabilidade Microbiana/efeitos da radiação , Permeabilidade , Radiação Ionizante , Reação em Cadeia da Polimerase em Tempo Real
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